ASPP2重组腺病毒通过调控NF-κB信号通路抑制DEN诱导的小鼠肝细胞癌发生  

ASPP2 recombinant adenovirus inhibits DEN-induced hepatocarcinogenesis in mice by regulating NF-κB signaling pathway

作　　者：高明慧 柴梦音[1] 寇卜心[1] 豆双双 刘晓霓[1] Gao Minghui;Chai Mengyin;Kou Buxin(Beijing Institute of Liver Diseases,You'an Hospital,Capital Medical University,Beijing 100069,China)

机构地区：[1]首都医科大学附属北京佑安医院北京肝病研究所,北京市

出　　处：《实用肝脏病杂志》2024年第2期169-173,共5页Journal of Practical Hepatology

基　　金：北京市自然科学基金资助项目(编号:7192084);首都卫生发展科研专项基金资助项目(编号:2020-2-1152);北京市公共医学研究所发展改革试点研究项目(编号:京医研2021-10)。

摘　　要：目的研究ASPP2重组腺病毒(ASPP2-ad)对二乙基亚硝胺(DEN)诱导的小鼠肝细胞癌(HCC)发生的抑制作用及其可能的作用机制。方法采用DEN腹腔注射联合0.005%DEN饮水方法构建小鼠HCC模型。将动物分为DEN处理组和DEN-ASPP2-ad处理组,每组10只。使用小动物超声成像系统动态观察小鼠HCC形成情况,大体记数肝脏肿瘤数量,采用免疫组化法检测肿瘤组织Ki67表达,采用流式多重蛋白定量技术检测小鼠血清IL-1β、IL-6、KC、IL-2和TNFα水平,采用Western blot法检测组织AFP、caspase3、cyclinD1、PCNA、p-IKK、IKK、p-IκB、IκB、p-p65和p65蛋白表达,采用实时定量PCR法检测Nfatc1 mRNA水平。结果DEN诱导24周后,DEN组小鼠肝脏肿瘤数为(9.9±1.9)个,显著多于DEN联合ASPP2-ad组[(3.9±1.2)个,P<0.05];DEN组小鼠Ki67阳性细胞数为(91.4±9.1)个,显著多于DEN联合ASPP2-ad组[(56.6±10.5)个,P<0.05];在实验40周末,DEN组小鼠生存率为65.2%,显著低于DEN联合ASPP2-ad组的90.0%(P<0.05);DEN组小鼠血清ALT和AST水平分别为(271.5±143.8)U/L和(299.3±221.4)U/L,均显著高于DEN-ASPP2-ad干预组[分别为(101.7±44.6)U/L和(124.1±75.0)U/L,P<0.05];DEN联合ASPP2-ad组血清IL-6和TNFα水平分别为(8.1±1.6)MFI和(8.1±1.0)MFI,均显著低于DEN组[分别为(16.3±0.4)MFI和(26.3±5.3)MFI,P<0.05];DEN/ASPP2-ad处理组AFP、Cyclin D1和PCNA表达减弱,而caspase-3表达增强,NF-κB信号通路蛋白(p-IKK、p-IκB和p-p65)表达减弱,p-IKK/IKKα、p-IκB/IκB和p-p65/p65比值也降低,NF-κB下游癌基因Nfatc1表达减弱(P<0.05)。结论ASPP2-ad可能通过调控NF-κB通路显著抑制DEN诱导的炎性增殖反应,阻抑HCC的发生,值得深入研究。Objective The aim of this experiment was to investigate the inhibitory effect of apoptosis stimulating protein 2 of P53(ASPP2)recombinant adenovirus(ASPP2-ad)on hepatocellular carcinogenesis induced by diethylnitrosamine(DEN)in mice.Methods The mouse liver cancer model was established by intraperitoneal injection of DEN at dose of 25 mg·kg-1 and drinking water with 0.005%of DEN.The experiment was divided into two groups with 10 mice in each group,receiving DEN or DEN and ASPP2-ad combination intervention.The formation of liver cancer in mice was observed by ultrasonography.The number of liver tumors was recorded,and the expression of Ki67 was detected by immunohistochemistry.Serum IL-1β,IL-6,KC,IL-2 and TNFαlevels were detected by flow multiprotein quantification technology.The protein expressions of AFP,caspase3,cyclin D1,PCNA,p-IKK,IKK,p-IκB,IκB,p-p65 and p65 were observed by Western blot,and the Nfatc1 mRNA level was assayed by real-time quantitative PCR.Results After 24 weeks of DEN induction,the number of liver tumors in DEN-intervened group was(9.9±1.9),significantly greater than[(3.9±1.2),P<0.05]in the DEN and ASPP2-ad combination intervened group;the number of Ki67 positive cells in DEN-intervened group was(91.4±9.1),significantly greater than[(56.6±10.5),P<0.05]in DEN/ASPP2-ad-intervened group;the survival rate in the DEN-intervened group was 65.2%,significantly lower than 90.0%(P<0.05)in DEN/ASPP2-ad-intervend group(P<0.05);serum ALT and AST levels were(271.5±143.8)U/L and(299.3±221.4)U/L,both significantly higher than[(101.7±44.6)U/L and(124.1±75.0)U/L,P<0.05]in DEN/ASPP2-ad-intervend group;serum IL-6 and TNFαlevels in DEN/ASPP2-ad-intervend group were(8.1±1.6)MFI and(8.1±1.0)MFI,both much lower than[(16.3±0.4)MFI and(26.3±5.3)MFI,P<0.05]in DEN-intervened group;the cancerous AFP,Cyclin D1 and PCNA expression became weaker,the p-IKK,p-IκB and p-p65 expression down-regulated,the p-IKK/IKKα,p-IκB/IκB and p-p65/p65 ratio down-regulated,the Nfatc1 expression weaker,while the caspase-3

关 键 词：肝细胞癌 P53凋亡刺激蛋白2 二乙基亚硝胺 核因子ΚB 小鼠 

分 类 号：R735.7[医药卫生—肿瘤]